1. Field of the Invention
The present invention relates to a particle measuring apparatus, in particular, to a particle measuring apparatus capable of distinguishing target particles in a sample from similar non-target particles for detection and counting. The apparatus is useful in, for example, accurately detecting and counting platelets and aggregations of platelets.
2. Description of the Related Art
Hitherto, counting of the number of particles, such as erythrocytes, leukocytes or platelets, or analysis of the particles has been carried out by obtaining an electrical or optical characteristic parameter with a flow cytometer.
A platelet function test is conducted to test a drop or a rise in the function of platelets. The platelet function test includes various tests such as a platelet adhesion test and a platelet releasing test. Among them, a platelet aggregation test has been frequently carried out, which is for inspecting aggregation of platelets.
In the platelet aggregation test, there has been hitherto used an absorbance method of measuring the degree of platelet aggregation in platelet rich plasma (PRP) as a change in absorbance (Tsukada Toshiyasu, PLATELET FUNCTION TEST AND PROBLEMS THEREOF, Japan Internal Medicine Journal, Vol. 80, No. 6, pp. 822-827, Jun. 10, 1991). This absorbance method comprises adding an aggregation-inducing agent into a given amount of the PRP and measuring the degree of light-shielding (absorbance) caused by aggregations of platelets. It has been used to detect a relatively large aggregation (in which about several hundreds of a single platelet are aggregated).
On the other hand, there are cases where the number of platelets is below several ten thousands/.mu.L in sudden thrombocytopenia or the like. Also, the number of platelets decreases to a non-measurable level at the time of chemotherapy or bone marrow transplant. When the number of platelets decreases, it is necessary to perform platelet transfusion or administration of hematopoietics. As a judgment standard for the above treatment, the number of platelets is used. Thus, it is extremely important to obtain the number of platelets accurately, especially when the number of platelets is at a low level.
The factors for causing the decrease in the measurement accuracy of the number of platelets may be, for example, particle components such as dusts, bacteria and air bubbles, or appearance of electric noises. These factors will be hereafter referred to as "background noise". For example, particle components may be mingled into a sheath liquid immediately after exchanging sheath liquids. These "background noise" may possibly generate signals similar to signals from platelets and are erroneously counted to show an untrue, high value about the number of platelets.
With respect to the platelet aggregation test, it is known in clinical medicine that platelet function rises in thrombotic diseases such as arteriosclerosis, hyperliperia, diabetes and hypertension, which are adult diseases. It is desired that a so-called thrombus preparing state, which is the state before turning to a thrombus, is discovered at an early stage.
In this thrombus preparing state, a feeble aggregation of platelets (aggregation of several platelets) appear at the initial stage, and consequently a measuring method for detecting feeble aggregations is necessary. In developing an anti-platelet agent or in a safety test for developing a certain medicine, it is desired to detect a feeble aggregation.
In using an anti-coagulation agent, aggregations of platelets may also appear depending on a specimen. Appearance of platelet aggregations makes it impossible to count the number of platelets accurately, and results in an untrue, low or high value about the number of platelets.
According to prior flow cytometers, light is applied to a fine sample stream containing target particles so as to measure information of light from each particle (for example, the intensity of forward scattered light and the intensity of fluorescence), thereby counting the number of the object particles and analyzing them.
For example, a two-dimensional scattergram (two-parameter scattergram) is prepared, using the obtained forward scattered light intensity (FSC) and fluorescence intensity (FL) as characteristic parameters, and then various analyzing treatments are conducted on the basis of the scattergram to separate particle groups from each other and to count the number of particles. For example, an erythrocyte and a platelet are different from each other in their sizes and shapes and consequently they can be separated into clearly different clusters on the scattergram.
A reticulated platelet has a relatively high RNA content among platelets. It is known that the number of reticulated platelets decreases in the case of diseases such as aplastic anemia and acute myelogeneous leukemia. Accordingly, in the case of blood diseases, it is considered to be useful as an indicator of platelet creation in the bone marrow that reticulated platelets are counted and analyzed.
In such a manner as above, the number of platelets can be counted by use of the two-dimensional scattergram (two-parameter scattergram) prepared by means of a flow cytometer. It has been found, by a measuring apparatus (FIC) for capturing images of particles in a sample stream which is made very fine by the flow cytometer, that particles different from platelets, such as fragmented erythrocytes, are present near the cluster of platelets obtained on the two-dimensional scattergram (two-parameter scattergram). A fragmented erythrocyte is a fragment of an erythrocyte, i.e., a torn-off erythrocyte. When non-platelet particles similar to platelets, such as fragmented erythrocytes, are counted as platelets, the number of platelets cannot be calculated accurately. Also, it has been found that the platelet area on the two-dimensional scattergram (two-parameter scattergram) embraces fragmented leukocytes (including eucaryocytes). Also, this results in an error count of platelets.